eIF2A represses cell wall biogenesis gene expression in Saccharomyces cerevisiae.

Translation initiation is a complex and highly regulated process that represents an important mechanism, controlling gene expression. eIF2A was proposed as an alternative initiation factor, however, its role and biological targets remain to be discovered. To further gain insight into the function of eIF2A in Saccharomyces cerevisiae, we identified mRNAs associated with the eIF2A complex and showed that 24% of the most enriched mRNAs encode proteins related to cell wall biogenesis and maintenance. In agreement with this result, we showed that an eIF2A deletion sensitized cells to cell wall damage induced by calcofluor white. eIF2A overexpression led to a growth defect, correlated with decreased synthesis of several cell wall proteins. In contrast, no changes were observed in the transcriptome, suggesting that eIF2A controls the expression of cell wall-related proteins at a translational level. The biochemical characterization of the eIF2A complex revealed that it strongly interacts with the RNA binding protein, Ssd1, which is a negative translational regulator, controlling the expression of cell wall-related genes. Interestingly, eIF2A and Ssd1 bind several common mRNA targets and we found that the binding of eIF2A to some targets was mediated by Ssd1. Surprisingly, we further showed that eIF2A is physically and functionally associated with the exonuclease Xrn1 and other mRNA degradation factors, suggesting an additional level of regulation. Altogether, our results highlight new aspects of this complex and redundant fine-tuned regulation of proteins expression related to the cell wall, a structure required to maintain cell shape and rigidity, providing protection against harmful environmental stress.

Xrn1 biochemically associates with eisosome proteins after the post diauxic shift in yeast.

mRNA degradation is one of the main steps of gene expression, and a key player is the 5'-3' exonuclease Xrn1. In Saccharomyces cerevisiae , it was previously shown, by a microscopy approach, that Xrn1 is located to different cellular compartments, depending on physiological state. During exponential growth, Xrn1 is distributed in the cytoplasm, while it co-localizes with eisosomes after the post-diauxic shift (PDS). Here, we biochemically characterize the Xrn1-associated complexes in different cellular states. We demonstrate that, after PDS, Xrn1 but not the decapping nor Lsm1-7/Pat1 complexes associates with eisosomal proteins, strengthening the model that sequestration of Xrn1 in eisosomes preserves mRNAs from degradation during PDS.

A specialised SKI complex assists the cytoplasmic RNA exosome in the absence of direct association with ribosomes.

The Ski2-Ski3-Ski8 (SKI) complex assists the RNA exosome during the 3' to 5' degradation of cytoplasmic transcripts. Previous reports showed that the SKI complex is involved in the 3' to 5' degradation of mRNAs, including 3' untranslated regions (UTRs) and devoid of ribosomes. Paradoxically, we recently showed that the SKI complex directly interacts with ribosomes during the co-translational mRNA decay and that this interaction is necessary for its RNA degradation promoting activity. Here, we characterised a new SKI-associated factor, Ska1, that associates with a subpopulation of the SKI complex. We showed that Ska1 is specifically involved in the degradation of long 3'UTR-containing mRNAs, poorly translated mRNAs as well as other RNA regions not associated with ribosomes, such as cytoplasmic lncRNAs. We further show that the overexpression of SKA1 antagonises the SKI-ribosome association. We propose that the Ska1-SKI complex assists the cytoplasmic exosome in the absence of direct association of the SKI complex with ribosomes.

Paenibacillus faecis sp. nov., isolated from human faeces.

A spore-forming, rod-shaped Gram-strain-positive bacterium, strain 656.84T, was isolated from human faeces in 1984. It contained anteiso-C15 : 0 as the major cellular fatty acid, meso-diaminopimelic acid was found in the cell wall peptidoglycan, the polar lipid profile consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and aminophospholipids as the major components, and the predominant menaquinone was MK-7. The DNA G+C content was 52.9 mol%. The results of comparative 16S rRNA gene sequence studies placed strain 656.84T within the genus Paenibacillus. Its closest phylogenetic relatives were Paenibacillus barengoltzii and Paenibacillus timonensis. Levels of DNA-DNA relatedness between strain 656.84T and Paenibacillus timonensis CIP 108005T and Paenibacillus barengoltzii CIP 109354T were 17.3 % and 36.8 %, respectively, indicating that strain 656.84T represents a distinct species. On the basis of phenotypic and genotypic results, strain 656.84T is considered to represent a novel species within the genus Paenibacillus, for which the name Paenibacillus faecis sp. nov. is proposed; the type strain is 656.84T ( = DSM 23593T = CIP 101062T).

Assessment of DNA encapsulation, a new room-temperature DNA storage method.

A new procedure for room-temperature storage of DNA was evaluated whereby DNA samples from human tissue, bacteria, and plants were stored under an anoxic and anhydrous atmosphere in small glass vials fitted in stainless-steel, laser-sealed capsules (DNAshells(®)). Samples were stored in DNAshells(®) at room temperature for various periods of time to assess any degradation and compare it to frozen control samples and those stored in GenTegra™ tubes. The study included analysis of the effect of accelerated aging by using a high temperature (76°C) at 50% relative humidity. No detectable DNA degradation was seen in samples stored in DNAshells(®) at room temperature for 18 months. Polymerase chain reaction experiments, pulsed field gel electrophoresis, and amplified fragment length polymorphism analyses also demonstrated that the protective properties of DNAshells(®) are not affected by storage under extreme conditions (76°C, 50% humidity) for 30 hours, guaranteeing 100 years without DNA sample degradation. However, after 30 hours of storage at 76°C, it was necessary to include adjustments to the process in order to avoid DNA loss. Successful protection of DNA was obtained for 1 week and even 1 month of storage at high temperature by adding trehalose, which provides a protective matrix. This study demonstrates the many advantages of using DNAshells(®) for room-temperature storage, particularly in terms of long-term stability, safety, transport, and applications for molecular biology research.